scholarly journals Specificity of human tissue kallikrein towards substrates containing Phe–Phe pair of amino acids

1999 ◽  
Vol 339 (2) ◽  
pp. 473-479 ◽  
Author(s):  
Daniel C. PIMENTA ◽  
Julie CHAO ◽  
Lee CHAO ◽  
Maria A. JULIANO ◽  
Luiz JULIANO
Keyword(s):  
Amino Acids ◽  
Amino Acid ◽  
Human Tissue ◽  
Binding Protein ◽  
General Structure ◽  
Protein A ◽  
Tissue Kallikrein ◽  
Hybrid Peptides ◽  
Loop Sequence ◽  
Reactive Centre Loop

We have explored in detail the determinants of specificity for the hydrolysis by human tissue kallikrein (hK1) of substrates containing the Phe–Phe amino acid pair, after which hK1 cleaves kallistatin (human kallikrein-binding protein), a specific serpin for this protease, as well as somatostatin 1–14. Internally quenched fluorogenic peptides were synthesized with the general structure Abz-peptidyl-EDDnp [Abz, o-aminobenzoic acid; EDDnp, N-(2,4-dinitrophenyl)ethylenediamine], based on the natural reactive-centre loop sequence of kallistatin from P9 to P´13, and the kinetic parameters of their hydrolysis by hK1 were determined. All these peptides were cleaved after the Phe–Phe pair. For comparison, we have also examined peptides containing the reactive-centre loop sequences of human protein-C inhibitor (PCI) and rat kallikrein-binding protein, which were hydrolysed after Phe–Arg and Leu–Lys bonds, respectively. Hybrid peptides containing kallistatin–PCI sequences showed that the efficiency of hK1 activity on the peptides containing kallistatin and PCI sequences depended on both the nature of the P1 amino acid as well as on residues at the P- and P´-sides. Moreover, we have made systematic modifications on the hydrophobic pair Phe–Phe, and on Lys and Ile at the P3 and P4 positions according to the peptide substrate, Abz-AIKFFSRQ-EDDnp. All together, we concluded that tissue kallikrein was very effective on short substrates that are cleaved after the Phe–Arg pair; however, hydrolysis after Phe–Phe or other hydrophobic pairs of amino acids was more restrictive, requiring additional enzyme–substrate interaction and/or particular substrate conformations.

scholarly journals Hydrolysis of somatostatin by human tissue kallikrein after the amino acid pair Phe-Phe

1997 ◽  
Vol 327 (1) ◽  
pp. 27-30 ◽  
Author(s):  
Daniel C. PIMENTA ◽  
Maria A. JULIANO ◽  
Luiz JULIANO
Keyword(s):  
Amino Acids ◽  
Growth Hormone ◽  
Human Tissue ◽  
General Structure ◽  
Tissue Kallikrein ◽  
Growth Hormone Releasing Hormone ◽  
Amino Acid Pair ◽  
Reactive Centre Loop ◽  
Hydrolysis Of ◽  
Similar Kinetic

Somatostatin-(1–14) was hydrolysed by human tissue kallikrein at the Phe7-Trp8 bond, after a Phe-Phe pair of amino acids, with similar kinetic parameters to those described for human high- and low-molecular-mass kininogens. Substance P and human insulin, which also contain a Phe-Phe pair in their sequences, were both resistant. More details of this hydrolytic specificity of human tissue kallikrein were obtained by synthesizing and assaying internally quenched fluorescent peptides containing the sequence of somatostatin-(1–14), as well as the reactive-centre loop of human kallikrein-binding protein (kallistatin). We also observed that human tissue kallikrein hydrolysed growth hormone-releasing hormone at the Arg11-Lys12 bond, although this peptide contains in its structure a pair of leucines (Leu22-Leu23), in contrast with the Phe-Phe pair in somatostatin. We have also demonstrated the susceptibility to human tissue kallikrein of some chromogenic peptides with the general structure X-Phe-Phe-p-nitroanilide and of d-Pro-Phe-Phe-4-methylcoumaryl-7-amide.

scholarly journals Molecular cloning and characterization of the cDNA coding for C4b-binding protein, a regulatory protein of the classical pathway of the human complement system

1985 ◽  
Vol 230 (1) ◽  
pp. 133-141 ◽  
Author(s):  
L P Chung ◽  
D R Bentley ◽  
K B Reid
Keyword(s):  
Amino Acids ◽  
Amino Acid ◽  
Amino Acid Sequence ◽  
Cdna Sequence ◽  
Binding Protein ◽  
Regulatory Protein ◽  
Protein A ◽  
Classical Pathway ◽  
Amino Acid Residues ◽  
Cloned Cdna

By using synthetic oligonucleotides as probes, plasmid clones containing portions of cDNA coding for human C4b-binding protein were isolated from a liver cDNA library. The entire amino acid sequence of the C4b-binding protein can be predicted from this study of the cloned cDNA when allied to a previous sequence study at the protein level [Chung, Gagnon & Reid (1985) Mol. Immunol. 22, 427-435], in which over 55% of the amino acid sequence, including the N-terminal 62 residues, was obtained. The plasmid clones isolated allowed the unambiguous determination of 1717 nucleotides of cDNA sequence between the codon for the 32nd amino acid in the sequence of C4b-binding protein and the 164th nucleotide in the 3′ non-translated region. The sequence studies show that the secreted form of C4b-binding protein, found in plasma, is composed of chains of apparent Mr 70 000 that contains 549 amino acid residues. Examination of the protein and cDNA sequence results show that there are at least two polymorphic sites in the molecule. One is at position 44, which can be glutamine or threonine, and the other is at position 309, which can be tyrosine or histidine. Northern-blot analysis indicated that the mRNA for C4b-binding protein is approx. 2.5 kilobases long. The N-terminal 491 amino acids of C4b-binding protein can be divided into eight internal homologous regions, each approx. 60 amino acids long, which can be aligned by the presence in each region of four half-cystine, one tryptophan and several other conserved residues. These regions in C4b-binding protein are homologous with the three internal-homology regions that have been reported to be present within the Ba region of the complement enzyme factor B and also to the internal-homology regions found in the non-complement beta 2-glycoprotein I.

scholarly journals Specificity of S'1 and S'2 subsites of human tissue kallikrein using the reactive-centre loop of kallistatin: the importance of P'1 and P'2 positions in design of inhibitors

2003 ◽  
Vol 371 (3) ◽  
pp. 1021-1025 ◽  
Author(s):  
Daniel C. PIMENTA ◽  
Sandro E. FOGAÇA ◽  
Robson L. MELO ◽  
Luiz JULIANO ◽  
Maria A. JULIANO
Keyword(s):  
Human Tissue ◽  
Serine Proteases ◽  
Potent Inhibitor ◽  
Competitive Inhibition ◽  
Plasma Kallikrein ◽  
Tissue Kallikrein ◽  
Loop Sequence ◽  
Reactive Centre Loop ◽  
Hydrolysis Of ◽  
Peptide Analogues

We have demonstrated that the S´1 and S´2 subsites of human tissue kallikrein (hK1) play determinant roles in the recognition and hydrolysis of substrates. The presence of serine at position P´1 and arginine at P´2 resulted in the best substrate, Abz-Ala-Ile-Lys-Phe-Phe-Ser-Arg-Gln-EDDnp, which was derived from the kallistatin reactive-centre loop sequence and quencher groups o-aminobenzoic acid (Abz) and N-(2,4-dinitrophenyl)ethylenediamine (EDDnp). Serine and arginine are also the residues at positions P´1 and P´2 in human kininogen, from which hK1 releases Lys-bradykinin. Several peptide analogues of Abz-Ala-Ile-Lys-Phe-Phe-Ser-Arg-Gln-EDDnp, in which the Ser and Arg residues were substituted with various other amino acids, were synthesized and tested as substrates. Most of them were hydrolysed slowly, although they showed significant binding to hK1, as demonstrated by their competitive inhibition constants (Ki). Using this information, six peptides were designed, synthesized and assayed as inhibitors of hK1. Abz-Lys-Phe-Phe-Pro-Arg-Gln-EDDnp, Abz-Lys-Phe-Arg-Pro-Arg-Gln-EDDnp and acetyl-Lys-Phe-Phe-Pro-Leu-Glu-NH2 inhibited hK1 in the range 20–30 nM (letters in italics denote the d-form of the amino acid). The peptide acetyl-Lys-Phe-Phe-Pro-Leu-Glu-NH2 was a weak inhibitor for other serine proteases, as indicated by the higher Ki values compared with hK1, but this peptide was a potent inhibitor of human plasma kallikrein, which has a Ki value of 8 nM. This result was surprising, since this enzyme is known to be a restricted arginyl-hydrolase. In conclusion, acetyl-Lys-Phe-Phe-Pro-Leu-Glu-NH2 can be used as a leader compound to design specific inhibitors for hK1, plasma kallikrein, or for both at same time, if the inhibition of kinin release is the main goal.

Structural and functional studies on C4b-binding protein, a regulatory component of the human complement system

1985 ◽  
Vol 5 (10-11) ◽  
pp. 855-865 ◽  
Author(s):  
L. Ping Chung ◽  
Kenneth B. M. Reid
Keyword(s):  
Amino Acid ◽  
Binding Protein ◽  
Protein A ◽  
Limited Proteolysis ◽  
Binding Activity ◽  
Amino Acid Sequencing ◽  
Functional Studies ◽  
Before And After ◽  
Cofactor Activity ◽  
Long Chain

The binding and cofactor activities of C4b-binding protein were examined before and after limited proteolysis by pepsin, trypsin and chymotrypsin. The major fragments generated were characterized by amino acid sequencing, thus establishing the precise points of limited proteolysis. These studies allow a tentative assignment of the cofactor activity site to the residues 177–322 of the 549 amino acid long chain of C4b-binding protein but indicated that residues in the region 332–395 are important in the binding activity.

scholarly journals Optimization of food compositions according to the ideal protein profile

Food systems ◽  
2021 ◽  
Vol 4 (1) ◽  
pp. 4-11
Author(s):  
S. V. Zverev ◽  
V. I. Karpov ◽  
M. A. Nikitina
Keyword(s):  
Amino Acids ◽  
Amino Acid ◽  
Essential Amino Acid ◽  
Raw Materials ◽  
Protein Profile ◽  
Protein A ◽  
Sum Of Squares ◽  
Reference Protein ◽  
Ideal Protein ◽  
Amino Acid Score

The paper emphasizes the importance of not only the quantitative but also qualitative composition of protein in nutrition. The authors propose protein classification into three main groups according to the concept of reference (ideal) protein. A mathematical model is examined to solve the task of rational mixture production upon the given profile of reference protein. Two variants of the criterion for formation of optimal composition are described. One of them presents the classical sum of squares of the residual for essential amino acid scores and 1. The second also presents the sum of squares of the residual for essential amino acid scores and 1 but with regard to only those amino acids, which scores are less than 1. The minima of these criteria at the set of variants for the content of ingredients are taken as targeted functions. The algorithm and the program of calculation were realized in the program environment Builder C++ 6.0. The macro flowchart of the algorithm is presented and detailed description of each block is given. The program interface before and after the start of the calculation module is shown. The main windows and interpretation of the presented data are described. An example of realization of the proposed mathematical apparatus when calculating a food model composition is given. Plant components (white kidney beans, flax, peanut, grit “Poltavskaya», dry red carrot) were used as an object of the research. Most plant proteins were incomplete. It is possible to regulate the chemical composition including correction of a protein profile by combination of plant raw materials. Analysis of alternative variants demonstrated that minimum essential amino acid score in the first composition was 0.79 (by the first criterion), in the second 1.0 (by the second criterion); the reference protein proportion in the mixture was 10.8 and 13.5, respectively, according to the first and second criterion. The comparative results by other quality indicators for protein in the mixture are also presented: the coefficient of amino acid score difference (CAASD), biological value (BV), coefficient of utility, essential amino acids index (IEAA).

scholarly journals Isolation and Characterization of a Type I Gene Encoding a Light-harvesting Chlorophyll a/b-binding Protein of Photosystem II in Peach

1998 ◽  
Vol 123 (4) ◽  
pp. 493-499 ◽  
Author(s):  
Kyu H. Chung ◽  
Dennis E. Buetow ◽  
Schuyler S. Korban
Keyword(s):  
Amino Acids ◽  
Amino Acid ◽  
Photosystem Ii ◽  
Amino Acid Sequence ◽  
Woody Plants ◽  
Light Harvesting ◽  
Binding Protein ◽  
Transit Peptide ◽  
Type I ◽  
Polyadenylation Sites

A nuclear gene, Lhcb1*Pp1, encoding a light-harvesting chlorophyll a/b-binding protein of photosystem II has been isolated from peach [Prunus persica (L.) Batsch. `Stark Earliglo'] leaf genomic DNA, cloned, and sequenced. This gene encodes a precursor polypeptide of 267 amino acids with a transit peptide of 34 and a type I mature protein of 233 amino acids. The amino acid sequence of the mature polypeptide is 89% to 94% and 80% to 94% similar to those encoded by type I Lhcb genes of annual and other woody plants, respectively. In contrast, the amino acid sequence of the peach transit peptide is less conserved being 47% to 69% similar to those of annual plants and only 17% to 22% similar to those of other woody plants. The peach gene was used as a probe for Lhcb gene expression. Lhcb mRNA is detected in leaves of field-grown trees during June to October. Lhcb mRNA is detected at a high level in leaves of peach shoots grown in tissue culture in the light, but only at a trace level in leaves grown in the dark. Some Lhcb genes appear to be light-modulated in stems. Lhcb1*Ppl contains four potential polyadenylation sites. S1 nuclease analysis detected transcripts of the sizes expected from each of the four polyadenylation sites. All four are found in leaves of light-grown shoots and of field-grown trees throughout the growing season. In contrast, only three are detected in stems of light-grown shoots.

scholarly journals Specificity of human tissue kallikrein towards substrates containing Phe‒Phe pair of amino acids

1999 ◽  
Vol 339 (2) ◽  
pp. 473 ◽  
Author(s):  
Daniel C. PIMENTA ◽  
Julie CHAO ◽  
Lee CHAO ◽  
Maria A. JULIANO ◽  
Luiz JULIANO
Keyword(s):  
Amino Acids ◽  
Human Tissue ◽  
Tissue Kallikrein

scholarly journals Rhizobium leguminosarum Has a Second General Amino Acid Permease with Unusually Broad Substrate Specificity and High Similarity to Branched-Chain Amino Acid Transporters (Bra/LIV) of the ABC Family

2002 ◽  
Vol 184 (15) ◽  
pp. 4071-4080 ◽  
Author(s):  
A. H. F. Hosie ◽  
D. Allaway ◽  
C. S. Galloway ◽  
H. A. Dunsby ◽  
P. S. Poole
Keyword(s):  
Amino Acids ◽  
Amino Acid ◽  
Rhizobium Leguminosarum ◽  
Binding Protein ◽  
Amino Acid Transporters ◽  
Acid Uptake ◽  
Amino Acid Permease ◽  
Branched Chain Amino Acid ◽  
General Amino Acid Permease ◽  
Branched Chain

ABSTRACT Amino acid uptake by Rhizobium leguminosarum is dominated by two ABC transporters, the general amino acid permease (Aap) and the branched-chain amino acid permease (BraRl). Characterization of the solute specificity of BraRl shows it to be the second general amino acid permease of R. leguminosarum. Although BraRl has high sequence identity to members of the family of hydrophobic amino acid transporters (HAAT), it transports a broad range of solutes, including acidic and basic polar amino acids (l-glutamate, l-arginine, and l-histidine), in addition to neutral amino acids (l-alanine and l-leucine). While amino and carboxyl groups are required for transport, solutes do not have to be α-amino acids. Consistent with this, BraRl is the first ABC transporter to be shown to transport γ-aminobutyric acid (GABA). All previously identified bacterial GABA transporters are secondary carriers of the amino acid-polyamine-organocation (APC) superfamily. Also, transport by BraRl does not appear to be stereospecific as d amino acids cause significant inhibition of uptake of l-glutamate and l-leucine. Unlike all other solutes tested, l-alanine uptake is not dependent on solute binding protein BraCRl. Therefore, a second, unidentified solute binding protein may interact with the BraDEFGRl membrane complex during l-alanine uptake. Overall, the data indicate that BraRl is a general amino acid permease of the HAAT family. Furthermore, BraRl has the broadest solute specificity of any characterized bacterial amino acid transporter.

scholarly journals Characterization of a glutathione S-transferase and a related glutathione-binding protein from gill of the blue mussel, Mytilus edulis

1995 ◽  
Vol 305 (1) ◽  
pp. 145-150 ◽  
Author(s):  
P J Fitzpatrick ◽  
T O B Krag ◽  
P Højrup ◽  
D Sheehan
Keyword(s):  
Amino Acid ◽  
Mytilus Edulis ◽  
Blue Mussel ◽  
Binding Protein ◽  
Protein A ◽  
Gel Filtration ◽  
Cumene Hydroperoxide ◽  
Gill Tissue ◽  
Glutathione S Transferase ◽  
Sequencing Studies

The major isoenzyme of glutathione S-transferase (GST 1) was purified to homogeneity from cytosolic extracts of Mytilus edulis gill tissue by GSH-agarose affinity chromatography followed by Mono Q ion-exchange f.p.l.c. This enzyme was particularly active with 1-chloro-2,4-dinitrobenzene, ethacrynic acid and cumene hydroperoxide as substrates. Immunoblotting and amino acid sequencing studies indicate that the enzyme belongs to the Pi class of GSTs. A related protein which binds to GSH-agarose was also purified. This GSH-binding protein did not immunoblot with GST antisera and showed no detectable catalytic activity with GST substrates although its N-terminal sequence was similar to Mu-class GSTs. Gel-filtration chromatography indicated that GST 1 is a dimer and the GSH-binding protein a monomer. Mass spectrometry and SDS/PAGE indicate subunit molecular masses of 24 kDa (GST 1) and 25 kDa (GSH-binding protein), respectively. Both proteins have amino acid compositions typical of GSTs.

104 The Danish Perspective to Remove Medicinal Zinc and Reducing the Use of Antibiotics in Swine Production

2021 ◽  
Vol 99 (Supplement_1) ◽  
pp. 103-104
Author(s):  
Hanne Maribo
Keyword(s):  
Amino Acids ◽  
Amino Acid ◽  
Protein Level ◽  
Diet Composition ◽  
Raw Materials ◽  
Productivity Loss ◽  
Protein A ◽  
Swine Production ◽  
Trial Testing ◽  
Ideal Protein

Abstract Diarrhoea in weaners has been commonly controlled by adding medicinal zinc (2500 ppm), but by June 2022 this was no longer allowed. In Denmark, antibiotics are accepted for therapeutic use only and usage is registered on pen level and is monitored by Danish authorities. This increases the risk of post-weaning diarrhoea. SEGES has tested several tools, additives e.g. organic acids, diet composition, raw materials e.g. blood plasma. Lowering the protein level in the diet post-weaning is very efficient, but adversely affects productivity. The latest results show on average that a reduction in protein from 19% to 15% in the weaner diet (6-9kg) results in a 60% reduction in diarrhoea; however, it also leads to a productivity loss of 1-1,5 euro. Reducing the protein level from 19% to 16,5% reduces the frequency of diarrhoea by 30% and the productivity loss by approx. 0,3 euro. A trial testing the possibility for compensation for this loss in the weaner period by adding extra protein and amino acids in the finisher diet (30–115 kg) is running now and preliminary results will be presented. Further results from trials reducing diarrhoea by reducing protein, a new way to calculate ideal protein and amino acid balances as well as results from concept tests with weaners will be presented. Further new results evaluating ideal protein and amino acid balances will be presented.

Sign in / Sign up

Export Citation Format

Share Document